Thromboembolic diseases, especially the cardiovascular and cerebrovascular thromboembolic diseases, are common disease that seriously threaten the health of the population and cause high fatality and disability rate. Currently available anticoagulants, such as heparin (includes low molecular weight heparin (LMWH) such as enoxaparin), warfarin, platelet aggregation inhibitors (such as aspirin) and hirudin are already approved for clinical use, however, these anticoagulants increase the bleeding risk in patient. Bleeding is one of the most common and serious complications in the clinical anticoagulant and antithrombotic therapies, thus it is very important to develop novel anticoagulant and antithrombotic drugs with lower bleeding risk. The clinical data and a series of animal experimental studies indicate that fXIa (or fXI) should be a novel target for the protection against thrombosis with lower bleeding risk in recent years. So develop drugs target fXIa (or fXI) may be an alternative way to treat and prevent thromboembolic diseases with lower bleeding risk.
High level of factor XI is a risk factor for deep venous thrombosis (Meijers J C M et al. N Engl J Med. 2000; 342: 696-701), and fXI-deficient patients reduce the incidence of deep vein thrombosis and ischemic stroke (Salomon O et al. Thromb Haemost. 2011, 105:269-73; Salomon O et al. Blood. 2008, 111: 4113-4117). These data suggest that the deficiency or inhibition of fXI/fXIa are associated with protection against thrombosis. Moreover, human deficiency in FXI results in a rare bleeding disorder and show seldom spontaneous bleeding (Seligsohn U. J Thromb Haemost. 2009, 7 (suppl): 84-87). It suggests that the deficiency or inhibition of fXI/fXIa has minimal or no bleeding risk in human.
In animal models, the fXI knockout mice do not have excessive bleeding when challenged by surgical procedures, and they have normal hemostatic capacity but have significantly reduced venous and arterial thrombosis in response to various mechanical and chemical vessel injuries. FXI antibody effectively reduced thrombosis in the rats, rabbits and baboons, and had little effect on the bleeding time (Renné T et al. J Thromb Haemost. 2009, Suppl 1: 79-83). The fXI monoclonal antibody (aXIMab) reduced thrombus formation and thrombin generation in the baboon's artificial vessels, but had little or no effect on the bleeding time and platelet aggregation activity, therefore, blocking the fXI may offer therapeutic advantages over other antithrombotic agents in terms of bleeding complications (Tucker et al. Blood. 2009, 113: 936-944). Comparing with enoxaparin and warfarin, the antisense oligonucleotides (ASOs) for fXI effectively reduced venous thrombosis and arterial thrombosis without causing bleeding; ASOs increased the antithrombotic activity of the enoxaparin and clopidogrel but did not increase bleeding (Zhang H et al. Blood. 2010, 116 (22): 4684-4692), and was considered as a safer anticoagulant.
It's confirmed that the mice knocked out of tissue factor (TF), fVII, fV, fX and prothrombin can't live for a long time (Mackman N. Arterioscler Thromb Vasc Biol. 2005, 25: 2273-228), while fVIII−/− and fIX−/− deficient mice can survive, but show bleeding diathesis, which similar to the human hemophilia that caused by the deficiency of fVIII or fIX. However, fXI knockout mice (fXI−/− mice) can live healthy with normal hemostatic function. Moreover, The fXI knockout mice are protected from thrombus formation when compared to wild-type animals (Gailani D, Renné T. J Thromb Haemost 2007, 5: 1106-1112).
As mentioned above, the fXI deficiency and inhibition have been shown to be associated with lower bleeding risk. It means that the drugs target fXIa or fXI has advantages of lower bleeding risk compares with that target thrombin, fX, fVII, and other coagulation factors. At present, only a few candidates including fXI antibody, fXI antisense oligonucleotides, peptidomimetics, some small chemical molecules and a molecule in a sponge (Schumacher W A et al. Arterioscler Thromb Vasc Biol. 2010, 30 (3): 388-392) target fXIa have been found, while selective peptide inhibitor target fXIa still keep to be not found.
Hookworms are blood-feeding intestinal nematodes that can cause chronic gastrointestinal blood loss of their hosts. They secrete antihemostatic molecules to counteract host hemostatic responses and result in bleeding. We recently isolated a novel anticoagulant peptide, named AcaNAP10 from the hookworm Ancylostoma caninum, which can inhibit both fXIa and fVIIa/TF. Significantly, AcaNAP10 was the first anticoagulant that inhibits both fXIa and fVIIa/TF (Li D, et al. Biochem Biophys Res Commun. 2010, 392 (2):155-9).
In order to obtain specific inhibitors of fXIa, based on the amino acid sequence of AcaNAP10, we invented a series of anticoagulant polypeptides that can selectively inhibit fXIa. These anticoagulant polypeptides can be used for development of novel anticoagulant drugs to treat or prevent thromboembolic disease with lower bleeding risk.